在猪瘟病毒兔化弱毒疫苗株的5’非编码区设计一对引物和一条荧光探针，利用荧光定量PCR原理，结合 LightCycler检测系统，首次建立了定量检测猪瘟兔化弱毒苗方法。结果表明，该方法的灵敏度为l02拷贝数，线 性范围为lO 一lO ，达6个数量级；标准样品的变异系数为2-3％一5．1％ (n=10)，疫苗样品组内实验变异系数为 0．85％一2．8％ (n=5)、组间实验为2．5％一7．3％ (n=5)，对同一样品分5次RNA提取和逆转录，其变异系数为5．O％； 对9份疫苗样品进行了检测，与兔体定型热反应方法相比较，有很好的相关性；整个检测过程仅需4h。该法可望 取代传统的兔体定型热反应用于疫苗生产过程中的效价测定及指导疫苗的配制，也为猪瘟病毒分子生物学研究提 供了一种新的、简捷有效的工具。

A rapid an d reproducible method was first established for assessment of Hog cholera lapinized virus(HCLV)loads in Classical Swine Fever vaccine using Flurogenic quantitative PCR (FQ·PCR)combines LiIghtCycler sequence detection system．The method contains a pair of primers and an internal daul·labled fluorogenic probe spanning the part of 5’noncoding region(5’NCR)of HCLV, the use of such a probe combined with the 5’-3’nuclease activity of Taq polymerase allows direct quan tification of the PCR product by the detection of a fluorescent reporter released in th e course of the exponential phase of the PCR．Th e sensitivity of the assay was 10 copies per reaction．Th e assay is linear within 6-log dynamic rang．The coeficient of variation(CV)of the standard of Ct value is 2．3％ 一5．1％ (n=10)；The CV ofvaccine sample is 0．85％一2．8％ in intra—assay and 2．5％一7．3％ in inter·assay(n =5)，respectively；Th e CV of the same sam ple in diferent RNA isolation and reverse transcription iS 5．0 ％ (n=5)．Nine vaccines were quantified by this method and give similar but more accurate results compared to the conventional rabbit fever reaction．Th e entire assay,including RNA isolation，reverse tran scription，an d quan tification，could be completed within 4 hour s．In conclusion，the high sensitivity， simplicity,an d reproducibility of the HCLV RNA quan tification which allows the screening of large numbers of sam ples，combined with its wide dynamic ran g，makes this method especially suitable for evaluating th e viral loads an d guiding how to confect th e vaccine，it also provides a novel an d simple research tool for CSFV